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1.
Int J Mol Sci ; 25(16)2024 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-39201343

RESUMO

Blastocyst vitrification has significantly improved embryo transfer methods, leading to higher implantation success rates and better pregnancy outcomes in subsequent frozen embryo transfer cycles. This study aimed to simulate the transcriptional changes caused by vitrifying human blastocysts using mouse blastocysts as a model and to further investigate these changes' effects. Utilizing a human vitrification protocol, we implanted both vitrified and fresh embryos into mice. We observed the implantation success rates and performed transcriptomic analysis on the blastocysts. To validate the results from messenger RNA sequencing, we conducted reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) to measure the expression levels of specific genes. Based on mRNA profiling, we predicted the microRNAs responsible for the regulation and used qPCR basic microRNA assays for validation. Our observations revealed a higher implantation success rate for vitrified embryos than fresh embryos. Transcriptomic analysis showed that vitrified-warmed blastocysts exhibited differentially expressed genes (DEGs) primarily associated with thermogenesis, chemical carcinogenesis-reactive oxygen species, oxidative phosphorylation, immune response, and MAPK-related signaling pathways. RT-qPCR confirmed increased expression of genes such as Cdk6 and Nfat2, and decreased expression of genes such as Dkk3 and Mapk10. Additionally, gene-microRNA interaction predictions and microRNA expression analysis identified twelve microRNAs with expression patterns consistent with the predicted results, suggesting potential roles in uterine epithelial cell adhesion, trophectoderm development, invasive capacity, and immune responses. Our findings suggest that vitrification induces transcriptomic changes in mouse blastocysts, and even small changes in gene expression can enhance implantation success. These results highlight the importance of understanding the molecular mechanisms underlying vitrification to optimize embryo transfer techniques and improve pregnancy outcomes.


Assuntos
Blastocisto , Criopreservação , Implantação do Embrião , Perfilação da Expressão Gênica , MicroRNAs , Vitrificação , Animais , Blastocisto/metabolismo , Camundongos , Implantação do Embrião/genética , Feminino , Criopreservação/métodos , Perfilação da Expressão Gênica/métodos , Gravidez , MicroRNAs/genética , Transcriptoma , Transferência Embrionária/métodos , Regulação da Expressão Gênica no Desenvolvimento
2.
J Clin Med ; 11(7)2022 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-35407431

RESUMO

Background: To identify the correlation among female age, cellular aging markers, and aneuploidy rate in in vitro fertilization (IVF) and the preimplantation genetic test for aneuploidy (PGT-A) cycles. Methods: This is a prospective cohort study recruiting 110 infertile women between August 2017 and July 2018. They were divided into young-age (<38 years, n = 60) and advanced-age (≥38 years, n = 50) groups. Peripheral leukocytes were assessed, and the granulosa cells were pooled during oocyte pickup. Mitochondrial DNA (mtDNA) copy number and telomere length (TL) were measured using real-time polymerase chain reaction. PGT-A was performed on the NGS platform. Results: mtDNA copy number and TL were positively correlated in both leukocytes (rho = 0.477, p < 0.001) and granulosa cells (rho = 0.361, p < 0.001), but the two parameters in leukocytes were not correlated with those in granulosa cells. In the young-age group, TL in the granulosa cells was the only factor correlated with the aneuploidy rate (rho = −0.283, p = 0.044), whereas in the advanced-age group, age was the main factor (rho = 0.358, p = 0.018). Conclusions: TL in the granulosa cells was negatively correlated with the aneuploidy rate in the young-age group, supporting the application of PGT-A in younger women.

3.
Carcinogenesis ; 34(2): 475-85, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23172669

RESUMO

Chronic hepatitis B virus (HBV) infection is the major cause of hepatocellular carcinoma (HCC). The pre-S(2) mutant large HBV surface antigen (LHBS) in type II ground glass hepatocytes (GGHs) has been recognized as an emerging viral oncoprotein; it directly interacts with the c-Jun activation domain-binding protein 1 (JAB1) and subsequently causes hyperphosphorylation of the tumor-suppressor retinoblastoma and, consequently, leads to disturbed cell cycle progression. The interaction of the pre-S(2) mutant LHBS with JAB1 could provide a potential target for chemoprevention. In this study, we found that the preneoplastic type II GGHs showed a significant decrease of the cyclin-dependent kinase inhibitor p27(Kip1), which serves as a marker for pre-S(2) mutant-JAB1 complex formation. The histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA) elevated expression of the tumor-suppressor thioredoxin-binding protein 2 (TBP2), which subsequently enhanced the JAB1-TBP2 interaction and abolished the pre-S(2) mutant LHBS-induced degradation of p27(Kip1), which, in turn, recovered the normal cell cycle checkpoint. The pre-S(2) mutant LHBS-induced pro-oncogenic effects: increased cell proliferation, nuclear/cytoplasmic ratio and proliferating cell nuclear antigen expression, were all greatly ameliorated after SAHA treatments, which suggested SAHA as a promising chemopreventive agent for the pre-S(2) mutant oncoprotein-induced HCC. In conclusion, this study provides the mechanism of histone deacetylase (HDAC) inhibitor in preventing the pre-S(2) mutant-induced oncogenic phenotype. The HDAC inhibitor SAHA is therefore a potential chemopreventive agent for high-risk chronic HBV patients who may develop HCC.


Assuntos
Carcinoma Hepatocelular/prevenção & controle , Proliferação de Células/efeitos dos fármacos , Antígenos de Superfície da Hepatite B/metabolismo , Hepatite B Crônica/prevenção & controle , Ácidos Hidroxâmicos/farmacologia , Neoplasias Hepáticas/prevenção & controle , Mutação/genética , Precursores de Proteínas/metabolismo , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Complexo do Signalossomo COP9 , Carcinoma Hepatocelular/etiologia , Carcinoma Hepatocelular/metabolismo , Ciclo Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Inibidor de Quinase Dependente de Ciclina p27/genética , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Citoplasma/efeitos dos fármacos , Citoplasma/metabolismo , Feminino , Imunofluorescência , Antígenos de Superfície da Hepatite B/genética , Vírus da Hepatite B/efeitos dos fármacos , Hepatite B Crônica/complicações , Hepatite B Crônica/virologia , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Humanos , Técnicas Imunoenzimáticas , Imunoprecipitação , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Hepáticas/etiologia , Neoplasias Hepáticas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Peptídeo Hidrolases/genética , Peptídeo Hidrolases/metabolismo , Precursores de Proteínas/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Semelhantes à Proteína de Ligação a TATA-Box/genética , Proteínas Semelhantes à Proteína de Ligação a TATA-Box/metabolismo , Técnicas do Sistema de Duplo-Híbrido , Vorinostat
4.
J Biomed Sci ; 16: 84, 2009 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-19751529

RESUMO

BACKGROUND: Chronic hepatitis B virus (HBV) infection is an important cause of hepatocellular carcinoma (HCC) worldwide. The pre-S1 and -S2 mutant large HBV surface antigen (LHBS), in which the pre-S1 and -S2 regions of the LHBS gene are partially deleted, are highly associated with HBV-related HCC. METHODS: The pre-S region of the LHBS gene in two hundred and one HBV-positive serum samples was PCR-amplified and sequenced. A pre-S oligonucleotide gene chip was developed to efficiently detect pre-S deletions in chronic HBV carriers. Twenty serum samples from chronic HBV carriers were analyzed using the chip. RESULTS: The pre-S deletion rates were relatively low (7%) in the sera of patients with acute HBV infection. They gradually increased in periods of persistent HBV infection: pre-S mutation rates were 37% in chronic HBV carriers, and as high as 60% in HCC patients. The Pre-S Gene Chip offers a highly sensitive and specific method for pre-S deletion detection and is less expensive and more efficient (turnaround time 3 days) than DNA sequencing analysis. CONCLUSION: The pre-S1/2 mutants may emerge during the long-term persistence of the HBV genome in carriers and facilitate HCC development. Combined detection of pre-S mutations, other markers of HBV replication, and viral titers, offers a reliable predictive method for HCC risks in chronic HBV carriers.


Assuntos
Carcinoma Hepatocelular/epidemiologia , Análise Mutacional de DNA/instrumentação , DNA Viral/genética , Antígenos da Hepatite B/genética , Vírus da Hepatite B/genética , Hepatite B Crônica/virologia , Neoplasias Hepáticas/epidemiologia , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas do Envelope Viral/genética , Carcinoma Hepatocelular/etiologia , DNA Viral/sangue , Vírus da Hepatite B/patogenicidade , Hepatite B Crônica/sangue , Humanos , Neoplasias Hepáticas/etiologia , Mutação , Reação em Cadeia da Polimerase , Valor Preditivo dos Testes , Risco , Deleção de Sequência
5.
Biochem Biophys Res Commun ; 335(1): 181-7, 2005 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-16105547

RESUMO

HHR23A and hHR23B are the human homologs of Saccharomyces cerevisiae Rad23. hHR23B is associated with the nucleotide excision repair (NER) factor xeroderma pigmentosum C (XPC) protein and is required for global genome repair. The function of hHR23A is not yet clear. In this study, the potential function of the hHR23A protein was investigated using RNA interference techniques. The hHR23A knock-down (KD) construct diminished the RNA level of hHR23A protein by approximately 60%, and it did not interfere with expression of the hHR23B gene. Based on Southwestern immunoblot and host-cell reactivation assays, hHR23A(KD) cells were found to be deficient in DNA repair activity against the DNA damage caused by UVC irradiation. In these hHR23A(KD) cells, the XPC gene was not normally induced by UVC irradiation, indicating that the hHR23A protein is involved in NER through regulation of the DNA damage recognition protein XPC. Co-immunoprecipitation experiments revealed that hHR23A was associated with a small portion of hHR23B and the majority of p53 protein, indicating that hHR23A regulates the function of XPC by its association with the NER activator p53.


Assuntos
Reparo do DNA , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , DNA/genética , Proteínas de Saccharomyces cerevisiae/química , Linhagem Celular Tumoral , Sobrevivência Celular , Enzimas Reparadoras do DNA , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica/genética , Humanos , Ligação Proteica , Interferência de RNA , Proteína Supressora de Tumor p53/metabolismo
6.
Biochem Biophys Res Commun ; 305(4): 1109-15, 2003 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-12767947

RESUMO

Xeroderma pigmentosum C (XPC) is a DNA repair factor essential for global genome repair (GGR) in nucleotide excision repair (NER). In the present study we screened for factors regulated by XPC after DNA damage. Ultraviolet C (UVC) irradiation-induced stress response factors were analyzed by a cDNA microarray chip system in HeLa and XP4PA-SV xpc mutant cell lines. The principal component analysis (PCA) method was employed to identify groups of genes with similar expression patterns over time after UVC irradiation. The growth arrest and DNA damage-inducible gene gadd45, as well as a small group of other genes, was found to exhibit an inducible expression pattern after 30min of incubation in xpc mutants but not in HeLa cells. Subsequent studies showed that gadd45 gene expression post-UVC irradiation was also present in the GGR mutant cells xpa and xpd, but not in TCR mutant csb cells. This evidence indicates that gadd45 plays a regulatory role in GGR of NER.


Assuntos
Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica/efeitos da radiação , Biossíntese de Proteínas , Raios Ultravioleta , Linhagem Celular , Sobrevivência Celular/efeitos da radiação , Dano ao DNA , Reparo do DNA , Relação Dose-Resposta à Radiação , Fibroblastos/metabolismo , Fibroblastos/efeitos da radiação , Perfilação da Expressão Gênica , Células HeLa , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Mutação , Análise de Sequência com Séries de Oligonucleotídeos , Análise de Componente Principal , Proteínas/genética , Pele/citologia , Raios Ultravioleta/efeitos adversos , Proteínas GADD45
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